Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Reduction in the developmental potential of intrathymic T cell progenitors with age.

doi: 10.4049/jimmunol.173.1.245

Figure Lengend Snippet: FIGURE 4. Proliferation and survival of ETP from young and old mice. Thymocytes were freshly isolated from young and old mice and stained for ETP in combination with anti-Ki-67 or annexin V labeling to determine their proliferation or survival status, respectively. A, ETP were labeled with anti-Ki-67 or isotype control Abs following fixation and permeabilization. The upper panel shows representative data for single young and old ani- mals. The lower panel indicates data pooled from the indicated number of different aged mice at each time point. B, The level of apoptosis in ETP was evaluated by annexin V labeling. The upper panel shows representative data for single young and old animals. The lower panel indicates data pooled from the indicated number of different aged mice at each time point. Means and SDs are shown for n 2–6 animals as indicated. , p 0.02; , p 0.002; #, p 0.005; ##, p 0.0005.

Article Snippet: The frequency of apoptotic cells in freshly harvested thymocyte populations was assessed by labeling with annexin V using the Apoptosis Detection kit as described by the manufacturer (R&D Systems, Minneapolis, MN) in combination with Abs to various cell surface determinants.

Techniques: Isolation, Staining, Labeling, Control

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 1. FOXA2 expression is up-regulated in CRC patients. A,B) FOXA2 expression profile in CRC tissues from TCGA cohort. C) Representative IHC images for FOXA2 in CRC tissues and normal tissues (https://www.proteinatlas.org/). D) Kaplan-Meier analysis for relapse-free-survival (RFS) of CRC patients with high (n = 709) or low (n = 633) FOXA2 expression based on the median expression of FOXA2 from the Kaplan-Meier Plotter (https://kmplot.com/analysis/index.php?p = service). E) Images of IHC staining for FOXA2 in CRC tissues and the paired adjacent normal tissues from our cohort. Scale bar = 120 μm. F) IHC scores of FOXA2 expression levels in paired normal and CRC samples were quantified. G) RT-qPCR analysis for FOXA2 gene expression in human CRC tissues and the matched adjacent normal tissues (ANT) from our cohort (n = 60). H) FOXA2 protein expression in eighteen paired CRC tissues was examined using western blot. I) RT-qPCR (n = 5) and J) western blot (n = 4) assays for FOXA2 gene and protein expression levels in six CRC cell lines and non-tumor cell line NCM460. Data are marked as the means ± SD. *p < 0.05, **p < 0.01, ****p < 0.0001.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Expressing, Immunohistochemistry, Quantitative RT-PCR, Gene Expression, Western Blot

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 2. FOXA2 enhances the proliferation, migration and invasion of CRC cells in vitro or in vivo. CCK8 analysis for cell proliferation of HCT-116 and SW480 cells with FOXA2 A) knockdown or B) over-expression (n = 4). C) EdU staining of HCT-116 and SW480 cells transfected with sh-FOXA2 or oe-FOXA2 as shown (n = 4). Scale bar = 50 μm. D) Transwell analysis for CRC cell migration and invasion after FOXA2 knockdown or over-expression (n = 5). Scale bar = 50 μm. E) RT-qPCR analysis for EMT markers in HCT-116 and SW480 cells after FOXA2 knockdown or over-expression (n = 3). F) Tumor samples from the indicated groups of mice expressing the control vector, sh-FOXA2 or oe-FOXA2 plasmids (n = 5 or 4 in each). G) Tumor formation volume and H) tumor weights were measured. Data are marked as the means ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Migration, In Vitro, In Vivo, Knockdown, Over Expression, Staining, Transfection, Quantitative RT-PCR, Expressing, Control, Plasmid Preparation

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 3. Positive correlation between FOXA2 and Nrf2/GPX4 signaling in CRC cell lines. A) Positive correlation between FOXA2 expression and GPX4, NFE2L2 (Nrf2), NQO1, G6PD, and SLC7A11 in CRC patients from TCGA database. B) RT-qPCR analysis for genes including Nrf2, GCLC, NQO1, SOD1, GPX4, SLC7A11 and G6PD in HCT-116 and SW480 cells with FOXA2 knockdown or over-expression (n = 3). C,D) IF staining for GPX4 expression in CRC cell lines transfected with sh-FOXA2 or oe-FOXA2 (n = 4). Scale bar = 20 μm. E) Western blot analysis for GPX4, Nrf2, and NQO1 protein expression levels in FOXA2-diminished or -over-expressed HCT-116 and SW480 cells (n = 3). Data are marked as the means ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Expressing, Quantitative RT-PCR, Knockdown, Over Expression, Staining, Transfection, Western Blot

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 4. FOXA2 suppression induces ferroptosis in CRC cell lines. A,B) ROS production was measured by DCF-DA staining in CRC cells with FOXA2 knockdown or over-expression. Scale bar = 50 μm. C,D) Lipid ROS generation in HCT-116 and SW480 cells was measured by C11-BODIPY581/591 staining post-sh-FOXA2 or oe-FOXA2 plasmid transfection. Scale bar = 20 μm. E) MDA levels, (F) GSH contents and G) iron currents in the show groups of CRC cells were examined. H) HCT-116 and SW480 cells with FOXA2 knockdown were treated with ferroptosis inhibitor (Fer-1, 1 μM), necroptosis inhibitor (Nec-1, 10 μM), or apoptosis inhibitor (Z-VAD-FMK, 10 μM) for another 24 h. Then, all cells were collected for cell viability examination using CCK-8 analysis. Data are marked as the means ± SD (n = 5 in each). **p < 0.01, ***p < 0.001, ****p < 0.0001; ns, no significant difference.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Staining, Knockdown, Over Expression, Plasmid Preparation, Transfection, CCK-8 Assay

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 5. FOXA2 suppression sensitizes CRC cells to OXA treatment by facilitating ferroptosis. HCT-116 and SW480 cells were transfected with sh- FOXA2, followed by OXA treatments for another 24 h. Then, all cells were harvested for subsequent assays. A) CCK-8 analysis for cell viability evaluation (n = 4). B,C) DCF-DA staining was used to examine ROS production (n = 4). Scale bar = 50 μm. D,E) C11-BODIPY581/591 staining was conducted for the measurements of lipid ROS production (n = 4). Scale bar = 20 μm. F) MDA contents, G) GSH levels, and H) iron currents were assessed (n = 4). I) Ferroptosis hallmarks including Nrf2, NQO1, SOD1, GPX4 and SLC7A11 were calculated by RT-qPCR (n = 3). Data are marked as the means ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Transfection, CCK-8 Assay, Staining, Quantitative RT-PCR

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 6. FOXA2 knockdown induces ferroptosis in chemoresistant CRC cell lines. A) RT-qPCR analysis for Nrf2, NQO1, SOD1, GPX4 and SLC7A11 gene expression levels in drug-sensitive or -resistant HCT-116 and SW480 cells (n = 3). B) Western blot assay for GPX4, Nrf2, and NQO1 protein expression levels in HCT-116 and SW480 cells with or without chemoresistance (n = 4). C) Western blot assay for FOXA2, GPX4, Nrf2, and NQO1 protein expression levels in chemoresistant HCT-116 and SW480 cells with or without FOXA2 knockdown (n = 4). (D) GPX4 expression by IF staining in drug-resistant CRC cells after transfection with sh-FOXA2 (n = 4). Scale bar = 20 μm. E) ROS and (F) lipid ROS production by DCF-DA (Scale bar = 50 μm) and C11-BODIPY581/591 (Scale bar = 20 μm) staining, respectively, in chemoresistant HCT-116 and SW480 cells with FOXA2 knockdown (n = 4). G) MDA levels, H) GSH contents, and I) iron currents in drug-resistant HCT-116 and SW480 cells after FOXA2 knockdown (n = 4). Data are marked as the means ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Knockdown, Quantitative RT-PCR, Gene Expression, Western Blot, Expressing, Staining, Transfection

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 7. FOXA2 knockdown CRC cells are sensitive to OXA-induced ferroptosis via the Nrf2 activation. HCT-116 cells with FOXA2 knockdown or over- expression were incubated with OXA (10 μM) alone or combination with Nrf2 activator (ML334, 20 μM) or inhibitor (ML385, 5 μM) for an additional 24 h. SW480 cells co-transfected with sh-FOXA2 and Nrf2 plasmids, or oe-FOXA2 and si-Nrf2 were exposed to OXA (10 μM) treatment for another 24 h. Then, all HCT-116 and SW480 cells were harvested for studies as follows. A-H) DCF-DA (Scale bar = 50 μm) and C11-BODIPY581/591 (Scale bar = 20 μm) staining were performed to examine ROS and lipid ROS production in CRC cells treated as shown (n = 4). I,J) MDA levels in HCT-116 and SW480 cells were examined. K,L) Examination of iron currents in CRC cells. (M,N) Calculation for cellular MDA levels. O,P) Iron currents in CRC cells were assessed (n = 5). Q-T) Western blot analysis for Nrf2, NQO1, and GPX4 protein expression levels in CRC cells were conducted (n = 3). Data are marked as the means ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Knockdown, Activation Assay, Over Expression, Incubation, Transfection, Staining, Western Blot, Expressing

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 8. RSL3 promotes the sensitivity of FOXA2-overexpressed and chemoresistant CRC cells to OXA treatment. Drug-sensitive CRC cells with FOXA2 over-expression or chemoresistant CRC cells were subjected to OXA (10 μM), GPX4 inhibitor (RSL3, 100 nM) single or double incubation for another 24 h. Next, all cells were harvested for studies as following. A-D) Western blot and IF staining analysis for GPX4 protein expression levels in cells treated as shown (n = 3 or 4 in each). E,F) MDA levels and G,H) iron contents were examined (n = 4). I,J) Lipid ROS production was measured using C11- BODIPY581/591 staining (n = 4). K,L) Cell viability of the shown CRC cells with or without chemoresistance was examined using CCK-8 analysis (n = 4). Data are marked as the means ± SD. Scale bar = 20 μm. +p < 0.05, ++p < 0.01 versus the Ctrl group; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns, no significant difference.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Over Expression, Incubation, Western Blot, Staining, Expressing, CCK-8 Assay

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 9. Identifying a potent suppressor of FOXA2. A) Working model of the protocols used to identify potent binding proteins and suppressors of FOXA2. B) SDS-PAGE band showing the immunoprecipitated proteins in HCT-116 and HEK293T cells with anti-FOXA2 antibody vs the isotype-controlled IgG. C) Immunoblotting assay to validate the presence of FOXA2 in the immunoprecipitated samples. D) There were a total of 31 overlapping FOXA2 binding proteins identified both in HCT-116 and HEK293T cells, including the listed E3 ligases TRIM36, CHIP, TRIM25, RNF2, and MID1. E) Western blot indicating the FOXA2 expression in HCT-116 and HEK293T cells transfected with the shown plasmids. F) TRIM36 expression profile in CRC tissues from TCGA cohort. G) Correlation between FOXA2 and TRIM36 from TCGA database. H) TRIM36 protein expression in eighteen paired CRC tissues was examined using western blot. I) Spearman’s correlation between TRIM36 and FOXA2 protein expression in the eighteen CRC patients. J) Western blot analysis for TRIM36 and FOXA2 in CRC cells transfected with TRIM36 plasmids. (K) RT-qPCR analysis for FOXA2 in CRC cells with or without TRIM36 over-expression. L) FOXA2 protein expression levels in empty vector and TRIM36-overexpressed HCT-116 or SW480 cells were examined at the shown time after CHX (20 μg ml−1) incubation. M) Western blot indicating the influence of MG132 (10 μM) on FOXA2 expression levels in CRC cells with or without TRIM36 over-expression. Data are marked as the means ± SD. **p < 0.01, ****p < 0.0001.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Binding Assay, SDS Page, Immunoprecipitation, Western Blot, Expressing, Transfection, Quantitative RT-PCR, Over Expression, Plasmid Preparation, Incubation

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 10. TRIM36 interacts with FOXA2 and induces its K48-linked polyubiquitination. A) Co-IP analysis of HEK293T cells transfected with Flag-tagged FOXA2 and HA-tagged TRIM36. Anti-Flag and anti-HA antibodies were used for western blot assay. B) GST precipitation indicating the direct interaction of TRIM36 with FOXA2 using purified GST-FOXA2 and His-tagged TRIM36 (left) or purified GST-TRIM36 and His-FOXA2 (right) by western blot analysis. GST was defined as a control. C) IF images of HEK293T cells co-transfected with 24 h of Flag-tagged FOXA2 (red) and HA-tagged TRIM36 (green). Scale bar = 15 μm. D,E) Schematic indicating full-length and truncated TRIM36 (top) and FOXA2 (top) with representative Co-IP assays (bottom) for the mapping analysis of the domains responsible for the TRIM36/FOXA2 interaction in HEK293T cells. F) Lysates of HEK293T cells transfected with plasmids expressing HA-FOXA2, Myc-Ub and increasing amounts of Flag-TIRM36 were immunoprecipitated with anti-HA beads and immunoblotted using an anti-Myc antibody. G) Western blots showing FOXA2 ubiquitination in HEK293T cells transfected with the indicated plasmids in different combinations. ∆CC, deletion of the CC domain. H) Western blot analysis showing K48 ubiquitination of FOXA2 in HEK293T cells co-transfected with Flag-TRIM36 and Myc-K48-Ub or Myc-K63-Ub. I) Western blot analysis showing K48 ubiquitination of FOXA2 in HEK293T cells co-transfected with Flag-TRIM36, Flag-TRIM36 (ΔCC), and Myc-K48-Ub. (J) FOXA2 protein expression levels and its ubiquitination levels in 24 h of OXA-treated HCT-116 and SW480 cells.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Co-Immunoprecipitation Assay, Transfection, Western Blot, Control, Expressing, Immunoprecipitation, Ubiquitin Proteomics

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

doi: 10.1002/advs.202304521

Figure Lengend Snippet: Figure 11. Conditional knockout of FOXA2 in IECs ameliorates colitis-associated tumorigenesis in vivo. A) Scheme protocols of recombination in Villin- Cre; FOXA2f/f mice. B,C) RT-qPCR and western blot assays for FOXA2 gene and protein expression levels in colon crypts from the mice treated with or without AOM/DSS (n = 6). D) H&E and IHC staining of FOXA2 in colon from FOXA2f/f and FOXA2cKO mice (n = 8). Scale bar = 120 μm. E) Inflammation scores were quantified. (F) FOXA2 expression by IHC staining was calculated. G) Body weights of mice were recorded (n = 10–15 in each group). H) Overall survival rates for each group of mice (n = 10–15 in each group). I) Images for colons from all groups of mice. J) Colon length was measured (n = 15). (K) Tumor number on colon was examined (n = 15). L) IHC staining for KI-67 in colon tissues was performed (n = 5). Scale bar = 50 μm. Data are marked as the means ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns, no significant difference.

Article Snippet: The following primary antibodies were used: anti-FOXA2 antibody (#NBP202088, 1:1000), anti-NQO1 antibody (#NB200-209, 1:1000), and antiGPX4 antibody (#NBP2-75511, 1:1000) were obtained from Novus Biologicals (Littleton, CO, USA); anti-Nrf2 antibody (#12721, 1:1000), anti-KI-67 antibody (#34330, 1:1000), anti-Flag antibody (#14793), anti-HA antibody (#5017), anti-Myc antibody (#2276) and anti-GAPDH (#5174) were obtained from Cell Signaling Technology (Beverly, MA, USA); anti-Fibronectin antibody (#MA5-11981, 1:1000), and anti-TRIM36 antibody (#PA5-28401, 1:1000) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Techniques: Knock-Out, In Vivo, Quantitative RT-PCR, Western Blot, Expressing, Immunohistochemistry

Journal: Disease Markers

Article Title: Responsive Expression of MafF to β -Amyloid-Induced Oxidative Stress

doi: 10.1155/2020/8861358

Figure Lengend Snippet: Primer sequences used in q-PCR.

Article Snippet: The primary antibodies were as follows: rabbit anti-GCLC antibody (1 : 10000, Abcam, UK), rabbit anti-MafF antibody (1 : 1000, Proteintech Group, Rosemont, IL, USA), rabbit anti-MafG antibody (1 : 2000, Novus Biologicals, CO, USA), rabbit anti-MafK antibody (1 : 5000, Abcam), and mouse GAPDH monoclonal antibody (1 : 5000, Proteintech Group).

Techniques:

Journal: Disease Markers

Article Title: Responsive Expression of MafF to β -Amyloid-Induced Oxidative Stress

doi: 10.1155/2020/8861358

Figure Lengend Snippet: Analysis of sMaf expressions in the hippocampus of AD patients. The mRNA expressions of (a) MafF, (b) MafG, and (c) MafK in the hippocampus of AD patient databases. The mRNA expressions of (d) MafF, (e) MafG, and (f) MafK in the hippocampus of AD patients under Braak stages III-IV and V-VI. The correlations between (h) MafF, (i) MafG, and (j) MafK and GCLC in the hippocampus of AD patient databases. All data were presented as mean ± SEM. ∗ p < 0.05 and ∗∗ p < 0.01 versus the control group or the Braak stage III-IV group.

Article Snippet: The primary antibodies were as follows: rabbit anti-GCLC antibody (1 : 10000, Abcam, UK), rabbit anti-MafF antibody (1 : 1000, Proteintech Group, Rosemont, IL, USA), rabbit anti-MafG antibody (1 : 2000, Novus Biologicals, CO, USA), rabbit anti-MafK antibody (1 : 5000, Abcam), and mouse GAPDH monoclonal antibody (1 : 5000, Proteintech Group).

Techniques: Control

Journal: Disease Markers

Article Title: Responsive Expression of MafF to β -Amyloid-Induced Oxidative Stress

doi: 10.1155/2020/8861358

Figure Lengend Snippet: Constitutive expressions of sMafs in different tissues. The mRNA expressions of (a) MafF, (b) MafG, and (c) MafK in the brain (hippocampus, cerebral cortex, cerebellum, and brainstem), liver, lung, and kidney. The relative expressions of sMafs were normalized to GAPDH ( n = 3). All data were presented as mean ± SEM.

Article Snippet: The primary antibodies were as follows: rabbit anti-GCLC antibody (1 : 10000, Abcam, UK), rabbit anti-MafF antibody (1 : 1000, Proteintech Group, Rosemont, IL, USA), rabbit anti-MafG antibody (1 : 2000, Novus Biologicals, CO, USA), rabbit anti-MafK antibody (1 : 5000, Abcam), and mouse GAPDH monoclonal antibody (1 : 5000, Proteintech Group).

Techniques:

Journal: Disease Markers

Article Title: Responsive Expression of MafF to β -Amyloid-Induced Oxidative Stress

doi: 10.1155/2020/8861358

Figure Lengend Snippet: MafF expression was increased in response to A β -induced oxidative injury. The protein expressions of MafF, MafG, and MafK in the (a) hippocampus from APP/PS1 mice and in the (b) hippocampus from SD rats after A β injection and in (c) SH-SY5Y cells treated with A β (20 μ M, 48 h) ( n = 3–5). The relative expressions of proteins were normalized to GAPDH. All data were presented as mean ± SEM; ∗ p < 0.05 and ∗∗ p < 0.01 versus the C57 group, the sham group, or the control group.

Article Snippet: The primary antibodies were as follows: rabbit anti-GCLC antibody (1 : 10000, Abcam, UK), rabbit anti-MafF antibody (1 : 1000, Proteintech Group, Rosemont, IL, USA), rabbit anti-MafG antibody (1 : 2000, Novus Biologicals, CO, USA), rabbit anti-MafK antibody (1 : 5000, Abcam), and mouse GAPDH monoclonal antibody (1 : 5000, Proteintech Group).

Techniques: Expressing, Injection, Control

Journal: Disease Markers

Article Title: Responsive Expression of MafF to β -Amyloid-Induced Oxidative Stress

doi: 10.1155/2020/8861358

Figure Lengend Snippet: MafF expression participated in the oxidative stress caused by A β . The protein expressions of GCLC in the (a) hippocampus from APP/PS1 mice and in the (c) hippocampus from SD rats after A β injection and in (e) SH-SY5Y cells treated with A β (20 μ M, 48 h). The GSH levels in the (b) hippocampus from APP/PS1 mice and in the (d) hippocampus from SD rats after A β injection and in (f) SH-SY5Y cells treated with A β (20 μ M, 48 h). The protein expressions of MafF, MafG, and MafK in SH-SY5Y cells transfected with MafF siRNA for 48 h (g). GSH level in SH-SY5Y cells transfected with MafF siRNA (10 nM) for 48 h with or without A β (20 μ M) treatment (6 h) (h) ( n = 3–7). The relative expressions of proteins were normalized to GAPDH. All data were presented as mean ± SEM; ∗ p < 0.05 and ∗∗ p < 0.01 versus the C57 group, the sham group, the control group, or the NC group; # p < 0.05 versus the NC+A β group.

Article Snippet: The primary antibodies were as follows: rabbit anti-GCLC antibody (1 : 10000, Abcam, UK), rabbit anti-MafF antibody (1 : 1000, Proteintech Group, Rosemont, IL, USA), rabbit anti-MafG antibody (1 : 2000, Novus Biologicals, CO, USA), rabbit anti-MafK antibody (1 : 5000, Abcam), and mouse GAPDH monoclonal antibody (1 : 5000, Proteintech Group).

Techniques: Expressing, Injection, Transfection, Control

Journal: Disease Markers

Article Title: Responsive Expression of MafF to β -Amyloid-Induced Oxidative Stress

doi: 10.1155/2020/8861358

Figure Lengend Snippet: The expressions of sMafs in basal or under A β /H 2 O 2 treatment in different cell lines. The constitutive expressions of (a) MafF, (b) MafG, and (c) MafK in SH-SY5Y cells, HepG2 cells, and A549 cells. The protein levels of MafF, MafG, and MafK in (f) SH-SY5Y cells, (d, g) HepG2 cells, and (e, h) A549 cells treated with A β for 48 h or H 2 O 2 for 24 h ( n = 3–5). The relative expressions of sMafs were normalized to β -actin/GAPDH. All data were presented as mean ± SEM; ∗ p < 0.05 and ∗∗ p < 0.01 versus the control group.

Article Snippet: The primary antibodies were as follows: rabbit anti-GCLC antibody (1 : 10000, Abcam, UK), rabbit anti-MafF antibody (1 : 1000, Proteintech Group, Rosemont, IL, USA), rabbit anti-MafG antibody (1 : 2000, Novus Biologicals, CO, USA), rabbit anti-MafK antibody (1 : 5000, Abcam), and mouse GAPDH monoclonal antibody (1 : 5000, Proteintech Group).

Techniques: Control